Citation:

Pitzer, E., T. Beasley, G. Jung, K. McDaniel, W. Padgett, W. Winnik, AND D. Herr. Developmental emamectin benzoate exposure alters behavior and proteomics in Long Evans rats. Triangle Chapter Society for Neuroscience, Durham, NC, April 21, 2023.

Impact/Purpose:

New Approach Methodologies (NAMs) are being developed to foster assessment of Neurotoxicity and Developmental Neurotoxicity (DNT). Confidence in the use of these NAMs for risk decisions under FIFRA, TSCA and other authorities is increased when the outcomes in these assays are linked in a biologically plausible context to adverse outcomes of regulatory interest through Adverse Outcome Pathways (AOPs). This project aims to assess the unique protein signatures following developmental exposure to known neurotoxicants in vivo, as well as measure functional changes that can then be related to these protein level changes. These alterations in proteomics can then be applied to the AOP framework as biological key events. Ultimately these in vivo proteomic biomarkers will be compared to those observed in in vitro counterparts. Concordance of 'omics results from in vitro cell culture to ex vivo or in vivo experiments will help verify the predicitve validity of the DNT NAMs.

Description:

Early neurodevelopmental periods are vulnerable to environmental compounds. Unfortunately, there are few adverse outcome pathways (AOP) that explore these critical periods. AOPs detail a series of biological responses linked by key events following a molecular initiating event and ending in an adverse effect. We are using proteomic changes produced by xenobiotics that can occur during this susceptible period of development to assess molecular alterations which can be applied in the AOP framework. Additionally, behavioral endpoints were assessed to understand potential adverse outcomes that may be related to these molecular perturbations. Pregnant Long Evans rats were gavaged with emamectin benzoate (EB; 3.78 mg/kg in 5 mL/kg DI water) or vehicle, from gestational day 6 to postnatal day (PND) 21. For proteomic experiments, pup region specific brain tissues (cortex, hippocampus, striatum, hypothalamus, midbrain, brainstem, and cerebellum) were collected throughout the postnatal period of exposure (PND2, 8, 15, 22). Cortex samples were assessed for proteomic content using Orbitrap LC-MS and identified proteins were then further processed using Proteome Discover and Ingenuity Pathway Analysis software. Behaviors were assessed throughout the experimental period, starting as early as PND2, as well as throughout adulthood. Behavioral assays included pup righting (PND 2-7), a modified functional observational battery (PND 22, 41, and 63), locomotor activity in figure-eight mazes (PND 13, 17, 21, 29, and 64), novel object recognition (NOR; PND 34-37), acoustic startle response (PND 29 and PND 77), and the Morris water maze (MWM; PND 76-92). Proteomic analyses showed that protein signatures for EB-treated rats differed in the cortex by sex as well as by age. Analysis indicates alterations in proteins implicating presynaptic vesicle docking/fusion machinery, cell-cell adhesion receptors and proteins, and enzymes involved in transcription and/or translation regulation. Behaviorally we observed that EB-exposed offspring displayed altered ontogeny of locomotor activity at PND17, decreased startle response at PND 29, and uncoordinated hindlimb movements that began in early postnatal weeks and persisted into adulthood. Additional proteomic analysis of other brain regions and inclusion of cognitive neurobehavioral tasks, like MWM and NOR, are on-going.  Overall, we observe changes in proteomic signatures during developmental treatment with EB that differed across the postnatal period and between sexes. Behavioral changes are present in developmentally exposed EB rats, postnatally and through adulthood. The protein level changes and observed apical behavioral endpoints could aid in AOP development.  This is an abstract of a proposed presentation and does not necessarily reflect US EPA policy.

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